Using human mammary epithelial cells (HMECs), we performed a forward genetic screen for alterations that contribute to malignant transformation and identified a novel oncogene, named FAM83B. The N-terminus of FAM83B encodes a PLD motif, which is both necessary and sufficient for FAM83B-mediated transformation. Elevated PLD activity contributes to tumorigenesis through the increased production of phosphatidic acid (PA), a potent second messenger involved in regulating mTOR and MAPK signaling, both of which are significantly activated in FAM83B-expressing cells. Ablation of FAM83B from tumor-derived cells with elevated EGFR or HER2, or from HMECs transformed by activated RAS, inhibited their proliferation, anchorage-independent growth and tumorigenicity, supporting a role for FAM83B as a critical EGFR/RAS- signaling intermediary. Together with the analysis of FAM83B expression in human tumor specimens, which revealed its overexpression in a variety of human cancers, our results suggest that FAM83B represents a novel oncoprotein involved in upregulating MAPK and mTOR signaling. The goals of this proposal are to define the involvement of FAM83B in MAPK and mTOR signaling, and examine human cancer specimens to correlate FAM83B expression with the activation of these highly important signaling cascades. Our hypothesis is that the FAM83B-mediated production of PA results in the activation of RAF-MEK-ERK signaling and the PI3K-AKT-independent activation of mTOR signaling, which drives breast cancer development. Successful completion of the proposed studies will provide important information regarding whether FAM83B inhibition in cancer cells will simultaneously ablate MAPK and mTOR signaling, validating FAM83B as a future target for therapeutic intervention. The aims of this proposal are to (1) Determine the involvement of FAM83B in EGFR/RAS-mediated transformation and effector activation. (2) Define the role of FAM83B in mTOR activation, independently of the canonical PI3K-AKT signaling axis. (3) Determine whether elevation of FAM83B expression provides an alternative mechanism for hyperactivating MAPK and mTOR signaling in cancer. PUBLIC HEALTH RELEVANCE: Successful identification of novel proteins involved in HMEC transformation, such as FAM83B, is critical to expanding our understanding of the oncogene signaling interactions involved in driving breast epithelial hyperplasia. Novel proteins also provide opportunities for drug development that will expand the arsenal of therapies necessary for eradicating cancer and extending the life of patients.